Interactions between bile and pancreatic juice diversions on cholecystokinin release and pancreas in conscious rats

Pancreatic exocrine secretion in the conscious rat is regulated by proteases secreted by the pancreas, and cholecystokinin (CCK) is known to be involved in its mechanism. It has also been reported that the absence of either pancreatic juice or bile in the duodenum could stimulate pancreatic secretion. Therefore, differences in CCK release responding to the exclusion of bile, pancreatic juice (PJ), or both bile and pancreatic juice (BPJ) from the intestine were examined by using a bioassay for cholecystokinin. Plasma CCK levels were increased by all three treatments compared with the basal value, the order of their effects being BPJ greater than PJ greater than bile diversion, and CCK concentrations produced by BPJ diversion were much greater than can be explained as simply summed effect of exclusions of bile and PJ. Pancreatic exocrine secretions were significantly increased by PJ and BPJ diversions, but the effect of bile diversion on the pancreas was not statistically significant. An additional infusion of CR-1409 (0.1 mg/rat), one of CCK receptor antagonists, inhibited exocrine function stimulated by BPJ diversion. We conclude (i) BPJ diversion is the strongest endogenous stimulant on CCK release; (ii) the potentiation between bile and PJ diversions is induced on CCK release; (iii) pancreatic protein secretion during BPJ diversion is mainly modulated by CCK.     

Stimulative effect of a casein hydrolysate on exocrine pancreatic secretion that is independent of luminal trypsin inhibitory activity in rats.

We have previously demonstrated that proteins could stimulate pancreatic secretion independently of luminal bile-pancreatic juice (BPJ) in a BPJ-diverted rat. To determine whether luminal protease-independent pancreatic secretion occurs in normal rats with BPJ returned to the upper small intestine, we investigated the pancreatic secretory response to intraduodenal instillation of a casein hydrolysate or the synthetic trypsin inhibitor, FOY 305, at concentrations which could almost equally inhibit hydrolysis of the synthetic substrate for trypsin with the luminal content. FOY 305 at 10 micrograms/ml and casein hydrolysate solutions at both 100 and 200 mg/ml similarly inhibited approx. 80% of the tryptic activity in the luminal contents of the proximal small intestine. Intraduodenal administration of casein hydrolysate solutions (100 and 200 mg/ml) significantly increased pancreatic secretion in a dose-dependent manner. However, intraduodenal administration of FOY 305 (10 micrograms/ml) was ineffective for stimulating pancreatic secretion. These results demonstrate that dietary protein enhances pancreatic secretion independently of the masking of luminal trypsin activity in rats.     

Antihypertensive effect of tryptic hydrolysate of milk casein in spontaneously hypertensive rats

1. Repeated oral administrations of tryptic hydrolysate of bovine milk casein (CEI) showed antihypertensive effect in spontaneously hypertensive rats. 2. Single oral administration of CEI antagonized the pressor response to angiotensin I. 3. Bovine milk casein hydrolysate inhibited the angiotensin I-converting enzyme (ACE) activity. Three peptides with ACE-inhibiting activity were isolated from CEI. 4. It is suggested that ACE-inhibiting peptides in the tryptic hydrolysate milk casein are absorbed from the intestinal tract and produce an antihypertensive effect.     

Protein malnutrition and response of pancreatic acinar cells to stimulation by cholecystokinin

Pancreatic lobules were isolated from 2 groups of male Wistar rats after 23 days of diet. A control group (C) fed on a 20% protein diet (16% gluten + 4% casein) and an experimental group (E) on a 5% protein diet (4% gluten + 1% casein). After isolation, lobules were preincubated 10 min with 10 muCi [3H]-leucine, washed, then incubate within Krebs Ringer bicarbonate Hepes. Basal secretion, then stimulated secretion (50 pM of cholecystokinin (CCK] of radioactive and non-radioactive protein and amylase outputs were measured. During basal secretion, in (E) group, lobules secreted more proteins than (C) one, the same outputs of amylase and radioactive protein were observed in both groups. The stimulated secretion by CCK increased the outputs of non-radioactive protein and amylase of lobules (T) (2-3 fold), but was without effect on lobule (E) outputs. Therefore, a low-protein diet involved a decrease of CCK sensibility on acinar cells, this fact might be mediated by a decreasing number and/or affinity of their CCK receptors.     

Effect of adding albumin to solutions of cholecystokinin on pancreatic protein output and pancreatic polypeptide release

In four conscious dogs with chronic gastric and pancreatic Thomas fistulas we studied the effect of 99% pure cholecystokinin-33 (CCK-33) solutions on pancreatic secretion and PP release. CCK-33 was dissolved in 0.154 M NaCl alone or in the same solution containing 1 g per 100 ml dog albumin. The response of pancreatic protein output to increasing doses of CCK-33 (0.5, 1, 2, 4 IDU/kg per h) was significantly $\text{(P < 0.05)}$ higher when CCK was dissolved in NaCl with albumin than in NaCl alone. These results were confirmed by measuring CCK immunoreactivity in samples from tips of infusion lines by a gastrin radioimmunoassay. Release of pancreatic polypeptide (PP) following increasing doses of CCK-33 was also significantly $\text{(P < 0.05)}$ elevated when CCK was dissolved in an albumin-containing solution. There was a significant $\text{(P < 0.02)}$ correlation between plasma concentrations of PP and pancreatic protein output.This study suggests that albumin should be added to CCK-33 solutions to preserve biological activity. The biological effect of CCK-33 may be substantially underestimated if albumin is omitted.     

Effects of a casein hydrolysate prepared from Aspergillus oryzae protease on adjuvant arthritis in rats

We evaluated the effects of a casein hydrolysate (CH) prepared from Aspergillus oryzae protease on rat adjuvant arthritis, a model of human rheumatoid arthritis. CH was administered orally once a day to the animals for 22 d after the adjuvant injection. CH suppressed swelling in the adjuvant-uninjected hind paws, and a higher dose of CH suppressed the increase in arthritic score and swelling of the adjuvant-injected hind paws. A histopathological examination revealed evidence that the higher dose of CH suppressed the articular changes in the rats. In addition, CH suppressed the production of nitric oxide and prostaglandin E(2) in the plasma of the rats. These results suggest that CH had a suppressive effect on adjuvant arthritis by inhibiting the acute and chronic inflammatory reactions.     

Galanin inhibits rat pancreatic amylase release via cholinergic suppression.

The effects of galanin on pancreatic exocrine function were examined using rat pancreatic tissues. In anesthetized rats, galanin (40 micrograms/kg/h) decreased amylase secretion stimulated by 2-deoxy glucose (5.8 +/- 0.1 vs. 3.1 +/- 0.1 times basal) and cholecystokinin octapeptide (21.5 +/- 0.6 vs. 16.8 +/- 0.5), while not inhibiting bethanechol-stimulated secretion. In dispersed acini, there was no effect of galanin alone (10(-8) to 10(-13) M) on amylase release, nor did galanin (10(-6) or 10(-8) M) coincubation affect amylase release stimulated by bethanechol (10(-3) to 10(-7) M) or CCK-8 (10(-8) to 10(-13) M). Using pancreatic lobules, coincubation with galanin (10(-6) M) suppressed 75 mM KCl-stimulated amylase secretion and ACh release (10.1 +/- 0.6% vs. 7.3 +/- 0.4%). Veratridine-stimulated (10(-4) M) amylase secretion and ACh release (12.4 +/- 1.7% vs. 8.5 +/- 0.7%) were similarly diminished.     

Exocrine pancreatic secretion: effect of subarachnoidal application of cholecystokinin octapeptide in rats

Administration of cholecystokinin octapeptide (CCK-8) intravenously, or in the subarachnoidal surface of the olfactory lobe in rats, caused an increase in pancreatic protein and amylase secretion. It was observed that for subarachnoidal administration of CCK-8 both protein and amylase outputs were higher than that seen after i.v. injection. This result is consistent with the presence of central CCK receptors which when activated can enhance pancreatic exocrine secretion. The blockade of the effect of CCK by administration of CCK-8-specific antisera proves the specificity of the subarachnoidal CCK-8 stimulation.     

Effect of proglumide, a cholecystokinin receptor antagonist, on caerulein-stimulated pancreatic enzyme secretion and pancreatic polypeptide release in the dog

In five conscious dogs we studied the effect of proglumide, a cholecystokinin (CCK) antagonist, on caerulein-stimulated pancreatic secretion and release of pancreatic polypeptide (PP). Graded doses of caerulein (15-240 ng/kg per h) were infused intravenously. Experiments were repeated with a fixed infusion of proglumide (40 mg/kg per h). Release of PP following increasing doses of caerulein was significantly inhibited by proglumide (P less than 0.01). However, proglumide did not significantly affect caerulein-stimulated pancreatic protein secretion. Proglumide might be useful in defining the physiological role of CCK.     

Bombesin inhibits insulin release from isolated pancreatic islets of rats in vitro.

The effect of bombesin on insulin release from isolated pancreatic islets of rats was examined in vitro. Bombesin, at the doses ranging from 10 ng/ml to 1 microgram/ml, significantly inhibited 16.7 mM glucose-induced insulin release, while bombesin had no inhibitory effect on insulin release at 8.3 mM and 3.3 mM glucose. Moreover, bombesin also suppressed insulin release elicited by 10 mM arginine at the doses of 100 ng/ml and 1 microgram/ml. These results indicate that bombesin has a direct inhibitory action on insulin release.     

In vivo and in vitro effects of cholecystokinin octapeptide on the release of prolactin in rats

The effect of cholecystokinin octapeptide (CCK-8) on the release of prolactin (PRL) in rats was studied in vivo and in vitro. Intravenous injection of 5 micrograms/100 g BW of CCK-8 resulted in significant increase in the plasma PRL level after 10 and 20 min. CCK-8 at concentrations of 10(-11) M to 10(-7) M also caused dose-dependent stimulation of PRL release from dispersed cells of rat anterior pituitary. On the other hand, dopamine inhibited PRL release from dispersed cells of rat anterior pituitary in a dose-related manner at concentrations of 10(-8) M to 10(-6) M. Release of PRL from the cells was increased by addition of K+ at high concentration (53 mM) in a Ca++-dependent manner. Addition of 10(-3) M verapamil to the incubation medium inhibited CCK-8-induced PRL release from the cells. Addition of dopamine (10(-7) M) to the incubation medium inhibited PRL release from the cells induced by CCK-8 or high K+ (53 mM). These results indicate that CCK-8 acts directly on the anterior pituitary cells to stimulate PRL release and that calcium ion is involved in the mechanism of this effect.     

Regulation of cholecystokinin release from central nerve terminals.

The high abundance of the cholecystokinin octapeptide in various brain regions is expressed by involvement of this neuropeptide in diverse brain functions. This peptide is mostly, if not always, co-localized with classic transmitters in central nerve terminals. Since the functions of the coexisting transmitters are often different, differential regulation of their release is obvious. This differentiation is realized by differences in presynaptic localization, release dynamics, and calcium regulation. In addition, CCK release is locally modulated by receptors, kinases and phosphatases. The regulatory mechanisms of CCK release are placed into physiological perspective.     

In vivo and in vitro effects of cholecystokinin octapeptide on the release of growth hormone in rats

The effect of cholecystokinin octapeptide (CCK-8) on the release of growth hormone (GH) in rats was studied in vivo and in vitro. Intravenous injection of 5 micrograms/100 g BW of CCK-8 resulted in significant increase in the plasma GH level after 10 and 20 min. CCK-8 at concentrations of 10(-11)M to 10(-7)M also caused dose-dependent stimulation of GH release from dispersed cells of rat anterior pituitary. On the other hand, somatostatin (SRIF) inhibited GH release from dispersed cells of rat anterior pituitary in a dose-related manner at concentrations of 10(-7)M to 10(-9)M. Release of GH from the cells was increased by addition of K+ at high concentration (50 mM) in a Ca++-dependent manner. Addition of 10(-3)M verapamil to the incubation medium inhibited CCK-8-induced GH release from the cells. Addition of SRIF (10(-7)M) to the incubation medium inhibited GH release from the cells induced by CCK-8 or high K+ (50 mM). These results indicate that CCK-8 acts directly on the anterior pituitary cells to stimulate GH release and that calcium ion is involved in the mechanism of this effect.     

Purification of A-subtype pancreatic cholecystokinin receptor by immunoaffinity chromatography.

So far, no efficient affinity chromatography for CCK receptor purification has been reported that prevented obtention of sequenceable amounts of purified receptor. In this work, 10% of plasma membrane receptor sites were specifically cross-linked with the photoreactive cleavable agonist 125I-ASD-[Thr28, Ahx31]-CCK-25-33, solubilized by NP-40, chromatographied on immobilized wheat germ agglutinin and further immunopurified using anti-CCK antibodies to an overall rate of 3000-3600-fold. Analysis of eluted material demonstrated a protein migrating at Mr 85,000-100,000 and the absence of 35S-labeled impurity. This single and efficient affinity chromatography should provide enough homogeneous receptor protein for microsequence determination and leads to consider immunoaffinity chromatography on immobilized anti-ligand antibodies as a potential tool for purification of membrane receptors.     

Interaction of somatostatin and calcium in regulating insulin release from isolated pancreatic islets of rats.

The effect of synthetic somatostatin on insulin release was studied in vitro by using isolated islets of rats. Somatostatin, with concentrations from 10 ng/ml to 10μg/ml, inhibited insulin release induced by 16.7 mM glucose. Insulin release elicited by 10 μg/ml glucagon or 2 mM dibutyryl cyclic AMP was likewise inhibited by 100ng/ml somatostatin. By raising the calcium concentration of the incubation medium to 6 mM, glucose-induced insulin release was fully restored even in the presence of somatostatin.However, the same maneuver only partially counteracted the somatostatin inhibition of dibutyryl cyclic AMP-induced insulin release. These results suggest the involvement of calcium mobilization process in the inhibitory action of somatostatin.